GSM6809666: Empty vector-2; Nicotiana benthamiana; RNA-Seq
total RNA were extracted from two groups (p103:: MD08G1076200 /pBIN61-p19 (overexpression) and P103 (empty vector)/pBIN61-p19 (control)) of agroinfiltrated leaves. Four biological replicates were used for RNA-Seq. Each biological replicate was comprised of a pool of 3 plants with 4 agroinfiltrated leaves per plant. Total RNA extracts were obtained using the RNeasy plant mini kit (QIAGEN, Leusden, The Netherlands) coupled with on-column DNase I treatment, following the manufacturer's guidelines.  Total RNA integrity (RIN>8) and purity were assessed using a 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA, USA) and a Nanodrop ND1000 spectrophotometer (Thermo scientific, Villebon-sur-Yvette, France), respectively. Total RNA was quantified using a Qubit RNA assay kit (Life technologies, Carlsbad, CA, USA). cDNA libraries were prepared from 3 µg total RNA of each replicate according to the KAPA HyperPrep Kit manufacturer's guidelines (Roche, Basel, Switzerland). All libraries were analyzed and quantified as described in Legay et al. (2016). The pooled libraries were sequenced on an Illumina NextSeq500 (NextSeq 500/550 Mid Output Kit v2.5 (150 Cycles)) to generate 75 base-pairs paired-end reads.